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By maintaining the telomere lengths, telomerase can make the tumor cells avoid the apoptosis, thus, achieving the cell immortalization. In the past, a series of telomerase detection systems have been developed through utilizing the unique characteristic of telomerase extended primer. However, fluctuation of telomerase activity, along with the cell cycle progression, leads to ambiguous detection results. Here, we reported a dual signal output detection strategy based on cell-cycle synchronization for precisely detecting telomerase activities by using a new AIEgen probe SSNB. Experimental and simulating calculation results demonstrated that positively charged SSNB could interact with DNA through the electrostatic interaction and π-π interaction, as well as the hydrogen bonds. The aggregation of SSNB caused by the extended template strand primer (TP) could be observed in tumor cells, thus, indicating the telomerase activity in various cell lines. Furthermore, after cell cycle synchronization, it was found that the telomerase activity in the S phase was the highest, no matter from the fluorescence intensity or the ROS generation situation. Dual signal outputs of SSNB verified the significance and necessity of cell-cycle synchronization detection for telomerase activity. This strategy could open a new window for the biotargets of which activity is variational in time dimension.

Probe-modified nanopores/nanochannels are one of the most advanced sensors because the probes interact strongly with ions and targets in nanoconfinement and create a sensitive and selective ionic signal. Recently, ionic signals have been demonstrated to be sensitive to the probe-target interaction on the outer surface of nanopores/nanochannels, which can offer more open space for target recognition and signal conversion than nanoconfined cavities. To enhance the ionic signal, we investigated the effect of grafting density, a critical parameter of the sensing interface, of the probe on the outer surface of nanochannels on the change rate of the ionic signal before and after target recognition (β). Electroneutral peptide nucleic acids and negatively charged DNA are selected as probes and targets, respectively. The experimental results showed that when adding the same number of targets, the β value increased with the probe grafting density on the outer surface. A theoretical model with clearly defined physical properties of each probe and target has been established. Numerical simulations suggest that the decrease of the background current and the aggregation of targets at the mouth of nanochannels with increasing probe grafting density contribute to this enhancement. This work reveals the signal mechanism of probe-target recognition on the outer surface of nanochannels and suggests a general approach to the nanochannel/nanopore design leading to sensitivity improvement on the basis of relatively good selectivity.

The subcellular distribution of adenosine 5'-triphosphate (ATP) and the concentration of ATP in living cells dynamically fluctuate with time during different cell cycles. The dictated activation of the biosensing process in living cells enables the spatiotemporal target detection in single living cells. Herein, a kind of o-nitrobenzylphosphate ester hairpin nucleic acid was introduced as a photoresponsive DNA probe for light-activated ATP detection in single living cells. Two methods to spatiotemporally activate the probe in single living cells were discussed. One method was the usage of the micrometer-sized optical fiber (about 5 μm) to guide the UV light (λ = 365 nm) to selectively activate the photoresponsive DNA probe in single living cells. The second method involved a two-photon laser confocal scanning microscope to selectively irradiate the photoresponsive DNA probes confined in single living cells via two-photon irradiation (λ = 740 nm). ATP aptamer integrated in the activated DNA probes selectively interacted with the target ATP, resulting in dictated signal generation. Furthermore, the photoactivated biosensing process enables dictated dual-model ATP detection in single living cells with "Signal-ON" fluorescence signal and "Signal-OFF" electrochemical signal outputs. The developed photoactivated biosensor for dictated ATP detection with high spatiotemporal resolution in single living cells at a desired time and desired place suggests the possibility to monitor biomarkers during different cell cycles.

Hybridization chain reaction (HCR) amplification strategy has been extensively explored for the application of electrochemical DNA-based sensors. Despite the enhancement in its sensitivity using the HCR, such sensor platform exhibited significant sensor-to-sensor variations in current due to variations in probe counts and lengths. To circumvent this, we are developing here a calibration-free "O-N" approach to generate a ratiometric, unitless value that is independent of these variations. Specifically, this approach employs two types of redox reporters, denoted as "One reporter" and "N reporters", with the former attached on the capture DNA and the latter on H1 and H2 strands. By optimizing the attachment sites of these reporters onto DNA strands, we demonstrate a significantly enhanced sensitivity of such sensor platform by four orders of magnitude, achieving accurate, calibration-free measurement of nucleic acids including ctDNA directly in undiluted whole blood without the requirement to calibrate each individual sensor.

During the past few decades, bio-inspired nanochannels have been well developed and applied in biosensing, energy transfer, separation, and so on. Here, inspired by the synergistic effect of biological nanopores, biomimetic solid-state nanochannels with hydrophilic DNA probes at the inner wall (DNA@IW

The grafting density of probes at sensor interface plays a critical role in the performance of biochemical sensors. However, compared with macroscopic interface, the effects of probe grafting density at nanometric confinement are rarely studied due to the limitation of precise grafting density regulation and characterization at the nanoscale. Here, we investigate the effect from the grafting density of DNA probes on ionic signal for nucleic acid detection in a cylindrical nanochannel array (with diameter of 25 nm) by combing experiments and theories. We set up a theoretical model of charge distribution from close to inner wall of nanochannels at low probe grafting density to spreading in whole space at high probe grafting density. The theoretical results fit well with the experimental results. A reverse of ionic output from signal-off to signal-on occurs with increasing probe grafting density. Low probe grafting density offers a high current change ratio that is further enhanced using long-chain DNA probes or the electrolyte with a low salt concentration. This work develops an approach to enhance performance of nanochannel-based sensors and explore physicochemical properties in nanometric confines.

The determination of T4 polynucleotide kinase (PNK) activity and the screening of PNK inhibitors are critical to disease diagnosis and drug discovery. Numerous electrochemical strategies have been developed for the sensitive measurement of PNK activity and inhibition. However, they often suffer from additional labels and multiple steps of the detection process for the electrochemical readout. Herein, we have demonstrated an electrochemical DNA (E-DNA) sensor for the one-step detection of PNK with "signal-on" readout with no need for additional labels. In our design, the highly switchable double-stranded DNA (dsDNA) probes are immobilized on the gold nanoparticle-decorated molybdenum disulfide nanomaterial (MoS2-AuNPs), which possesses large surface area and high conductivity for elevating the signal gain in the PNK detection. This signal-on E-DNA sensor integrated with MoS2-AuNPs exhibits a much higher sensitivity than that without MoS2-AuNPs, showing a detection limit of 2.18 × 10-4 U/mL. Furthermore, this assay shows high selectivity, with the ability to discriminate PNK from other enzymes and proteins, and can be utilized to screen inhibitors. The proposed sensor is easy to operate with one-step readout and robust for PNK detection in the biological matrix and shows great potential for point-of-care in clinical diagnostics and drug screening.

The rational design of DNA capture probes for modulating the binding affinity to tune the dynamic range of electrochemical DNA (E-DNA) biosensors is valuable and effective. Most of current strategies, however, require designing several DNA capture probes to achieve the tunable dynamic range, which is cumbersome and costly. Herein, we develop the photoresponsive E-DNA biosensors with tunable dynamic ranges by using only one photocleavable capture probe (PC-CP). The photoresponsive PC-CP is a stem-loop DNA structure containing a photocleavable linker (PC-linker) in the loop. The PC-linker can be cleaved by UV irradiation to switch the structure of PC-CP, through which the binding affinity to the target could be tuned. In this way, the dynamic range, the sensitivity, and the specificity of photoresponsive E-DNA biosensors can be tuned. Furthermore, the developed photoresponsive E-DNA biosensors enable sensitive and selective detection of target DNA in complex samples with a tunable dynamic range, which offers the possibility of clinical applications.

DNA adenine methylation (Dam) MTase serves a very important epigenetic process that transfers a methyl group on an adenine residue including N6-methyladenosine (m6A). A variety of evidence have dem...

Using a surface forces apparatus (SFA), we have studied the nanomechanical behavior of short single-stranded and partially and fully double-stranded DNA molecules attached via one end to a self-assembled monolayer on a gold surface. Our results confirm the previously proposed "mushroom-like" polymer structure for surface-attached, single-stranded DNA at low packing density and a "brush-like" structure for the same construct at higher density. At low density we observe a transition to "rigid rod" behavior upon addition of DNA complementary to the surface-attached single strand as the fraction of molecules that are double-stranded increases, with a concomitant increase in the SFA-observed thickness of the monolayer and the characteristic length of the observed repulsive forces. At higher densities, in contrast, this transition is effectively eliminated, presumably because the single-stranded state is already extended in its "brush" state. Taken together, these studies offer insights into the structure and physics of surface-attached short DNAs, providing new guidance for the rational design of DNA-modified functional surfaces.

As a powerful tool for obtaining sufficient DNA from rare DNA resources, polymerase chain reaction (PCR) has been widely used in various fields, and the optimization of PCR is still in progress due to the dissatisfactory specificity, sensitivity and efficiency. Although many nanomaterials have been proven to be capable of optimizing PCR, their underlying mechanisms are still unclear. So far, the scientifically compelling and functionally evolving metal–organic framework (MOF) materials with high specific surface area, tunable pore sizes, alterable surface charges and favourable thermal conductivity have not been used for PCR optimization. In this study, UiO-66 and ZIF-8 were used to optimize error-prone two round PCR. The results demonstrated that UiO-66 and ZIF-8 not only enhanced the sensitivity and efficiency of the first round PCR, but also increased the specificity and efficiency of the second round PCR. Moreover, they could widen the annealing temperature range of the second round PCR. The interaction of DNA and Taq polymerase with MOFs may be the main reason. This work provided a candidate enhancer for PCR, deepened our understanding on the enhancement mechanisms of nano-PCR, and explored a new application field for MOFs., Many new materials have the ability to optimize polymerase chain reaction (PCR). Metal-organic frame materials UiO-66 and ZIF-8 can enhance sensitivity, specificity and efficiency of PCR, indicating their potential as PCR enhancers.

Gene therapy has immense potential as a therapeutic approach to serious diseases. However, efficient delivery and real-time tracking of gene therapeutic agents have not been solved well for successful gene-based therapeutics. Herein we present a versatile gene-delivery strategy for efficient and visualized delivery of therapeutic genes into the targeted nucleus. We developed an integrin-targeted, cell-permeable, and nucleocytoplasmic trafficking peptide-conjugated AIEgen named TD NCP for the efficient and sequential targeted delivery of an antisense single-stranded DNA oligonucleotide (ASO) and tracking of the delivery process into the nucleus. As compared with TD NCP/siRNA-NPs (siRNA functions mainly in the cytoplasm), TD NCP/ASO-NPs (ASO functions mainly in the nucleus) exhibited a better interference effect, which further indicates that TD NCP is a nucleus-targeting vector. Moreover, TD NCP/ASO-NPs showed a favorable tumor-suppressive effect in vivo.

DNA occupies significant roles in life processes, which include encoding the sequences of proteins and accurately transferring genetic information from generation to generation. Recent discoveries have demonstrated that a variety of biological functions are correlated with DNA's conformational transitions. The non-B form has attained great attention among the diverse forms of DNA over the past several years. The main reason for this is that a large number of studies have shown that the non-B form of DNA is associated with gross deletions, inversions, duplications, translocations as well as simple repeating sequences, which therefore causes human diseases. Consequently, the conformational transition of DNA between the B-form and the non-B form is important for biology. Conventional fluorescence probes based on the conformational transitions of DNA usually need a fluorophore and a quencher group, which suffers from the complex design of the structure and tedious synthetic procedures. Moreover, conventional fluorescence probes are subject to the aggregation-caused quenching (ACQ) effect, which limits their application toward imaging and analyte detection. Fluorogens exhibiting aggregation-induced emission (AIE) have attracted tremendous attention over the past decade. By taking advantage of this unique behavior, plenty of fluorescent switch-on probes without the incorporation of fluorescent quenchers/fluorophore pairs have been widely developed as biosensors for imaging a variety of analytes. Herein, the recent progress in bioanalytical applications on the basis of aggregation-induced emission luminogens (AIEgens)/nucleic acid nanostructures are presented and discussed.

Metal ions play a critical role in human health and abnormal levels are closely related to various diseases. Therefore, the detection of metal ions with high selectivity, sensitivity and accuracy is particularly important. This article highlights and comments on the coordination-induced structural changes of DNA-based optical, electrochemical and optical-electrochemical-combined sensors for metal ions detection. Challenges and potential solutions of DNA-based sensors for the simultaneous detection of multiple metal ions are also discussed for further development and exploitation.

Understanding the spatiotemporal effects of surface topographies and modulated stiffness and anisotropic stresses of hydrogels on cell growth remains a biophysical challenge. Here we introduce the photolithographic patterning or two-photon laser scanning confocal microscopy patterning of a series of o-nitrobenzylphosphate ester nucleic acid-based polyacrylamide hydrogel films generating periodically-spaced circular patterned domains surrounded by continuous hydrogel matrices. The patterning processes lead to guided modulated stiffness differences between the patterned domains and the surrounding hydrogel matrices, and to the selective functionalization of sub-regions of the films with nucleic acid anchoring tethers. HeLa cells are deposited on the circularly-shaped domains functionalized with the MUC-1 aptamers. Initiation of the hybridization chain reaction by nucleic acid tethers associated with the continuous hydrogel matrix results in stress-induced ordered orthogonal shape-changes on the patterned domains, leading to ordered shapes of cell aggregates bound to the patterns., Spatiotemporal patterning of hydrogel matrices has been used to control cell behavior. Here the authors present photoresponsive DNA-based hydrogels and demonstrate patterning of physical and biochemical properties to tune local cell responses.

The ability to monitor specific molecules in real-time directly in a flowing sample stream and in a manner that does not adulterate that stream could greatly augment quality control in, for example, food processing and pharmaceutical manufacturing. Because they are continuous, reagentless, and able to work directly in complex samples, electrochemical DNA-based (E-DNA) sensors, a modular and, thus, general sensing platform, are promising candidates to fill this role. In support, we describe here an E-DNA sensor supporting the continuous, real-time measurement of melamine in flowing milk. Using target-driven DNA triplex formation to generate an electrochemical output, the sensor responds to rising and falling melamine concentration in seconds without contaminating the product stream. The continuous, autonomous, real-time operation of sensors such as this could provide unprecedented safety, convenience, and cost-effectiveness relative to the batch processes historically employed in molecular quality control.

Dynamic nanoassembly-based drug delivery system (DNDDS) has evolved from being a mere curiosity to emerging as a promising strategy for high-performance diagnosis and/or therapy of various diseases. However, dynamic nano-bio interaction between DNDDS and biological systems remains poorly understood, which can be critical for precise spatiotemporal and functional control of DNDDS in vivo. To deepen the understanding for fine control over DNDDS, we aim to explore natural systems as the root of inspiration for researchers from various fields. This review highlights ingenious designs, nano-bio interactions, and controllable functionalities of state-of-the-art DNDDS under endogenous or exogenous stimuli, by learning from nature at the molecular, subcellular, and cellular levels. Furthermore, the assembly strategies and response mechanisms of tailor-made DNDDS based on the characteristics of various diseased microenvironments are intensively discussed. Finally, the current challenges and future perspectives of DNDDS are briefly commented.

The capability to image, as well as control and manipulate single molecules such as nucleic acids (DNA or RNA) can greatly enrich our knowledge of the roles of individual biomolecules in cellular processes and their behavior in native environments. Here we summarize the recent advances of single nucleic acid imaging based on optical observation and force manipulation. We start by discussing the superiority of single molecule image, the central roles nucleic acids play in biosystems, and the significance of single molecule image towards nucleic acids. We then list a series of representative examples in brief to illustrate how nucleic acid of various morphologies has been imaged from different aspects, and what can be learned from such characterizations. Finally, concluding remarks on parts of which should be improved and outlook are outlined.

Functional nucleic acids (FNAs)-based biosensors have shown great potential in heavy metal ions detection due to their low-cost and easy to operate merits. However, in most FNAs based fluorescence probes, the ingenious designs of double-labeled (fluorophore and quencher group) DNA sequence, not only bring the annoyance of organic synthesis, but also restrict its use as a robust biosensor in practical duties. In this paper, we design a simple AIEgens functional nucleic acids (AFNAs) probe which consists of only fluorogen but no quencher group. With the help of duplex-specific nuclease (DSN) enzyme based target recycling, high fluorescence signal and superior sensitivity towards Hg2+ are achieved. This robust assay allows for sensitive and selective detection of Hg2+ in real water samples and mapping of intracellular Hg2+, without double-labeling of oligonucleotide with a dye-quencher pair, nor the multiple assay steps.

Fluorophore probes are widely used for bioimaging in cells, tissues, and animals as well as for monitoring of multiple biological processes in complex environments. Such imaging properties allow scientists to make direct visualizations of pathological events and cellular targets. Conventional fluorescent molecules have been developed for several decades and achieved great successes, but their emissions are often weakened or quenched at high concentrations that might suffer from the aggregation-caused quenching (ACQ) effect, which reduces the efficiencies of their applications. In contrast to the ACQ effect, aggregation-induced emission (AIE) luminogens (AIEgens) display much higher fluorescence in aggregated states and possess various advantages such as low background, long-term tracking ability, and strong resistance to photobleaching. Therefore, AIEgens are employed as unique fluorescence molecules and building blocks for biosensing applications in the fields of ions, amino acids, carbohydrates, DNAs/RNAs, peptides/proteins, cellular organelles, cancer cells, bacteria, and so on. Quite a few of the above biosensing missions are accomplished by modular peptide-modified AIEgen probes (MPAPs) or modular DNA-modified AIEgen probes (MDAPs) because of the multiple capabilities of peptide and DNA modules, including solubility, biocompatibility, and recognition. Meanwhile, both electrostatic interactions and coupling reactions could provide efficient methods to construct different MPAPs and MDAPs, finally resulting in a large variety of biosensing probes. Those probes exhibit leading features of detecting nucleic acids or proteins and imaging mass biomolecules. For example, under modular design, peptide modules possessing versatile recognition abilities enable MPAPs to detect numerous targets, such as integrin αvβ3, aminopeptidase N, MMP-2, MPO, H2O2, and so forth; MDAP could allow the imaging of mRNA in cells and tissue chips, suggesting the diagnostic functions of MDAP in clinical samples. Modular design offers a novel strategy to generate AIEgen-based probes and expedites functional biomacromolecules research. In this vein, here we review the progress on MPAPs and MDAPs in the most recent 10 years and highlight the modular design strategy as well as their advanced biosensing applications including briefly two aspects: (1) detection and (2) imaging. By the use of MPAPs/MDAPs, multiple bioanalytes can be efficiently analyzed at low concentrations and directly visualized through high-contrast and luminous imaging. Compared with MPAPs, the quantities of MDAPs are limited because of the difficult synthesis of long-length DNA strands. In future work, multifunctional of DNA sequences are needed to explore varieties of MDAPs for diverse biosensing purposes. At the end of this Account, some deficiencies and challenges are mentioned for briging more attention to accelerate the development of AIEgen-based probes.

The spatially defined functionalization of microparticles with asymmetric shape-controlled nucleic acid patterns is a major challenge in materials science. The asymmetric patterning of microparticles is important to allow the controlled fabrication of crystalline lattices or controlled aggregates of microparticles. We present the combination of two-photon lithography and photocleavable o-nitrobenzylphosphate ester nucleic acid coating-modified microparticles as a versatile means to asymmetrically pattern single microparticle surfaces. The two-photon patterning of microparticles with predesigned nucleic acid structures of different sizes (700 nm to 2.8 μm) and shapes (circles, rings, triangles, and squares) are demonstrated. In addition, complex patterned domains consisting of two different asymmetric nucleic acid domains are fabricated by the controlled Z-positioning of the microparticles in respect to the two-photon irradiation sources. In addition, the two-photon lithographic patterning of the photocleavable DNA coating allows the generation of functional nucleic acid domains for the photostimulated activation of the catalytic hybridization assembly (CHA) of branched nucleic acid structures on single microparticles.

Current visual biosensing methods, including colorimetric-based, fluorescence-based and chemiluminescence-based methods, are inappropriate for the hundreds of millions of people affected by color blindness and color weakness. Compared with these available methods, a droplet motion-based strategy might be a promising protocol for extension to a wider user base. Here we report a protocol for manipulating the hydrophobicity of DNA, which offers a droplet motion-based biosensing platform for the visual detection of small molecules (ATP), nucleic acids (microRNA) and proteins (thrombin). The protocol starts with target-triggered rolling-circle amplification that can readily generate short single-stranded DNA (ssDNA) fragments or long ssDNA. By exploiting macroscopic wetting behavior and molecular interaction, one can tailor the conformation of ssDNA on the water-oil interface to control the relevant DNA hydrophobicity. The wettability of DNA can be translated into visual signals via reading the sliding speed or the critical sliding angle. The time range for the entire protocol is ∼1 d, and the detection process takes ∼1 min.

The fabrication of sensitive sensors with high selectivity is highly desirable for the detection of some important biomarkers, such as nucleic acids, proteins, small molecules and ions. DNA hybridization chain reaction (HCR) and DNA supersandwich self-assembly (SSA) are two prevalent enzyme-free signal amplification strategies to improve sensitivity of the sensors. In this review, we firstly describe the characteristics about DNA HCR and DNA SSA, and then summarize the advances in the one-dimensional DNA nanostructures assisted by HCR and SSA. This review has been divided into three parts according to the two signal amplification methods and highlights recent progress in these two strategies to improve the detection sensitivity of proteins, nucleic acids, small molecules and ions.

Single nucleotide polymorphisms (SNPs) are the most abundant genetic polymorphisms and are responsible for many genetic diseases and cancers. In general, SNPs detection is performed by a single probe system (SPS), in which a single probe specifically hybridizes to one target. However, with the use of this method it is hard to improve the hybridization specificity and single mismatched discrimination factors (DF). In addition, the multiprobe system (MPS) requires complex probe designs and introduces at least one auxiliary probe except for the probe complementary to the target, resulting in a complicated detection system. Faced with these difficulties, we perform the SNP detection using a d/l-tryptophan (Trp) guided DNA probe and regulate the DF of electrochemical DNA (E-DNA) sensors by molecular chirality. We show that the DF of the d-Trp incubated E-DNA sensor (d-sensor) is larger than that of the l-sensor. More importantly, we achieve the high specificity by coupling d-Trp and l-Trp incubated E-DNA sensors, and the median DF is 7.21. Furthermore, the specificity of SNP detection can be further improved by supersandwich assay, and the median DF is enlarged to 37.23, which is comparable to that obtained with a multiprobe detection system.

Absorption of ultraviolet (UV) light by nucleic acid could lead to mutations and skin cancers. Traditional damage detection methods based on fluorescence not only need dye/quencher groups but also display relatively high background interference, causing difficulty in synthesis and purification and thus low specificity of detection. Here, by combining rolling circle amplification (RCA) and aggregation-induced emission molecules (AIE), we made up for the defects of traditional methods to some extent and could also differentiate damaged and undamaged DNA. We also studied radiation damage of the p53 gene fragment both from UVA and UVC, although the mechanism of UVA in mutagenesis remains controversial. To amplify the signal-to-background ratio, we ligated the linear p53 (L p53) gene fragment to be a circular p53 (C p53) gene fragment, which is a key component for RCA. The combination of RCA products and positive TPE-Z (quaternized tetraphenylethene salt) molecules induced the aggregation of AIE molecules, and subsequently resulted in significant fluorescence enhancement (the signal for the undamaged DNA is 598% higher than that of the damaged). Compared with the traditional aggregation-caused quenching (ACQ) based fluorescent method, our assay was more sensitive and more specific.

The advances in bioinformatics and biomedicine have promoted the development of biomedical imaging and theranostic systems to respectively extend the endogenous biomarker imaging with high contrast and enhance the therapeutic effect with high efficiency. The emergence of biomacromolecule-functionalized aggregation-induced emitters (AIEgens), utilizing AIEgens, and biomacromolecules (nucleic acids, peptides, glycans, and lipids), displays specific targeting ability to cancer cell, improved biocompatibility, reduced toxicity, enhanced therapeutic effect, and so forth. This review summarizes the rational design of biomacromolecule-functionalized AIEgens and their biomedical applications in recent ten years, including high-resolution optical imaging of cell, tissue, and small animal model with low background; the biomarker detection for early diagnosis and prognosis; the delivery and monitoring of prodrugs; image-guide photodynamic therapy and its combination with chemotherapy. Through illustrating their functional mechanisms and application, it is hoped that this review would open up a completely new train of research thought for attracted researchers in various fields.

Two aerobic and obligately acidophilic bacteria, designated HZ411T and 4MSH08T, were isolated from the forest soil of Dinghushan Biosphere Reserve, Guangdong Province, PR China (23° 10′ N, 112° 31′ E). These two strains were Gram-stain-negative short rods that multiplied by binary division. HZ411T was motile, with a single polar flagellum, but 4MSH08T was non-motile. The results of the 16S rRNA gene sequence analysis indicated that these two strains formed a common clade with members of the genus Edaphobacter in subdivision 1 of the phylum Acidobacteria but they each occupied a unique position in the genus. HZ411T and 4MSH08T had a 16S rRNA gene sequence similarity of 96.2 % between each other and the highest sequence similarity of 97.7 and 96.9 % to Edaphobacter modestus Jbg-1T and Edaphobacter aggregans Wbg-1T, respectively. The DNA–DNA hybridization rate between HZ411T and E. modestus Jbg-1T was 22.7 %. The DNA G+C contents of HZ411T and 4MSH08T were 57.7 and 59.3 %, respectively. HZ411T and 4MSH08T had similar major (>10 %) fatty acids with very high percentages of iso-C15 : 0 and C16 : 1ω7c, and similar major polar lipid profiles (both contained a phosphatidylethanolamine, phosphatidyldimethylethanolamine and glycolipid and several unidentified aminolipids and polar lipids). On the basis of these physiological, phylogenetic and chemotaxonomic data, we suggest that HZ411T and 4MSH08T represent two novel species of the genus Edaphobacter , for which the names Edaphobacter flagellatus HZ411T (=GDMCC 1.1193=LMG 30085) and Edaphobacter bradus 4MSH08T (=GDMCC 1.1317=KCTC 62475) are proposed.

Detection of an ultralow concentration of mRNA is important in the prognosis of gene-related diseases. In this study, a DNA-conjugated amphiphilic aggregation-induced emission probe (TPE-R-DNA) was synthesized for cancer tissue imaging and prognosis analysis based on an exonuclease III-aided target recycling technique. TPE-R-DNA comprise two components: a hydrophobic component that serves as the “turn-on” long wavelength fluorescence imaging agent (TPE-R-N3); and a hydrophilic single DNA strand (Alk-DNA) which acts as specific recognition part for target mRNA. In the absence of target mRNA, TPE-R-DNA had almost no fluorescence because of its high water solubility. Conversely, the TPE-R-DNA was digested by exonuclease III (Exo III) in the presence of MnSOD mRNA to release the hydrophobic fluorogens (TPE-R-AT). Subsequently, TPE-R-AT formed aggregates, and therefore, fluorescence signal was distinctly observed. For the first time, the structure of the hydrolysis product (TPE-R-AT), containing two bases A an...

Although extensively used in all fields of chemistry, molecular recognition still suffers from a significant limitation: host-guest binding displays a fixed, hyperbolic dose-response curve, which limits its usefulness in many applications. Here we take advantage of the high programmability of DNA chemistry and propose a universal strategy to engineer biorecognition-based sensors with dual programmable dynamic ranges. Using DNA aptamers as our model recognition element and electrochemistry as our readout signal, we first designed a dual signaling "signal-on" and "signal-off" adenosine triphosphate (ATP) sensor composed of a ferrocene-labeled ATP aptamer in complex to a complementary, electrode-bound, methylene-blue labeled DNA. Using this simple "dimeric" sensor, we show that we can easily (1) tune the dynamic range of this dual-signaling sensor through base mutations on the electrode-bound DNA, (2) extend the dynamic range of this sensor by 2 orders of magnitude by using a combination of electrode-bound strands with varying affinity for the aptamers, (3) create an ultrasensitive dual signaling sensor by employing a sequestration strategy in which a nonsignaling, high affinity "depletant" DNA aptamer is added to the sensor surface, and (4) engineer a sensor that simultaneously provides extended and ultrasensitive readouts. These strategies, applicable to a wide range of biosensors and chemical systems, should broaden the application of molecular recognition in various fields of chemistry.

As a biomarker for early cancer diagnosis, telomerase are one of the promising targets for cancer therapeutics. Inspired by the fluorescent emission principle of aggregation-induced emission fluorogens, we creatively designed an AIE-based turn-on method to detect telomerase activity from cell extracts. A positively charged fluorogen (TPE-Z) is not fluorescent when freely diffused in solution. The fluorescence of TPE-Z is enhanced with the elongation of the DNA strand which could light up telomere elongation process. By exploitation of it, we can detect telomerase activity from different cell lines (E-J, HeLa, MCF-7, and HLF) with high sensitivity and specificity. Moreover, our method is successfully employed to demonstrate the applications in bladder cancer diagnosis (41 urine specimens from bladder cancer patients and 15 urine specimens from normal people are detected). The AIE-based method provides a simple one-pot technique for quantification and monitoring of the telomerase activity and shows great potential for future use in clinical tests.

Nature owns remarkable capabilities in sensing target molecules, while the artificial biosensor lags far behind nature. Inspired by nature, we devise a new sensing platform that can specifically bind the molecules and synchronously initiate a specific signal response. We rationally designed a type of bipolar probe that is comprised of a hydrophilic DNA part and a hydrophobic conjugated polymer (CP) unit. In aqueous solution, they can form micelles with a hydrophobic CP core and a hydrophilic DNA shell. The aggregation-caused quenching suppresses the fluorescence of CP. Adding telomerase, the hydropathical profile of the bipolar probes is drastically regulated that results in the collapse of micelles and liberates fluorescence simultaneously. The probe has been used in both mimic systems and real urine samples (38 samples). We achieve sensitive and specific detection of telomerase and obtain clearly classification for normal people and cancer patients. It can also be used in a signal off sensor that is used to detect mercury ions.

3D cross-linked DNA superstructures switch off the ionic flux through solid-state nanopores with extremely high ON-OFF ratios of 10(3) -10(5) . This gating mechanism can be generally applicable in a wide range of nanopores with opening diameters up to 650 nm. The 3D bio-supramolecular gatekeepers outperform previous low-dimensional or simple-structured DNA functional components.

The enzyme-free toehold-mediated strand displacement reaction has shown potential for building programmable DNA circuits, biosensors, molecular machines and chemical reaction networks. Here we report a simple colorimetric method using gold nanoparticles as signal generators for the real-time detection of the product of the strand displacement cascade. During the process the assembled gold nanoparticles can be separated, resulting in a color change of the solution. This assay can also be applied in complex mixtures, fetal bovine serum, and to detect single-base mismatches. These results suggest that this method could be of general utility to monitor more complex enzyme-free strand displacement reaction-based programmable systems or for further low-cost diagnostic applications.

ZIF-8(zinc-methylimidazolate framework-8), one of the zeolitic imidazolate frameworks (ZIFs), as quenching platforms for detection of HIV-1 DNA, which was identified to be effective for highly sensitive detection of HIV-1 DNA. The enhanced fluorescence signal has a relationship with the ssDNA concentration, the detection limit is as low as 1.2 nmol L-1 with good selectivity. This study is an important step toward rational design of materials to achieve specific interactions between biomolecules and synthetic particle surfaces.

The concatenated hybridization chain reaction (C-HCR) was constructed as a versatile and robust tool for signal amplification and intracellular imaging, which was attributed to the synergistic amplification effect between HCR-1 and HCR-2., Biomolecular self-assembly has spurred substantial research efforts for the development of low-cost point-of-care diagnostics. Herein, we introduce an isothermal enzyme-free concatenated hybridization chain reaction (C-HCR), in which the output of the upstream hybridization chain reaction (HCR-1) layer acts as an intermediate input to activate the downstream hybridization chain reaction (HCR-2) layer. The initiator motivates HCR-1 through the autonomous cross-opening of two functional DNA hairpins, yielding polymeric dsDNA nanowires composed of numerous tandem triggers T as output of the primary sensing event. The reconstituted amplicon T then initiates HCR-2 and transduces the analyte recognition into an amplified readout, originating from the synergistic effect between HCR-1 and HCR-2 layers. Native gel electrophoresis, atom force microscopy (AFM) and fluorescence spectra revealed that C-HCR mediated the formation of frond-like branched dsDNA nanowires and the generation of an amplified FRET signal. As a versatile and robust amplification strategy, the unpreceded C-HCR can discriminate DNA analyte from its mutants with high accuracy and specificity. By incorporating an auxiliary sensing module, the integrated C-HCR amplifier was further adapted for highly sensitive and selective detection of microRNA (miRNA), as a result of the hierarchical and sequential hybridization chain reactions, in human serum and even living cells through an easy-to-integrate “plug-and-play” procedure. In addition, the C-HCR amplifier was successfully implemented for intracellular miRNA imaging by acquiring an accurate and precise signal localization inside living cells, which was especially suitable for the ex situ and in situ amplified detection of trace amounts of analyte. The C-HCR amplification provides a comprehensive and smart toolbox for highly sensitive detection of various biomarkers and thus should hold great promise in clinical diagnosis and assessment. The infinite layer of multilayered C-HCR is anticipated to further strengthen the amplification capacity and reliability (anti-invasion performance) of intracellular imaging approach, which is of great significance for its bioanalytical applications.

The specific recognition and programmable assembly properties make DNA a potential material for nanodevices. However, the more intelligent the nanodevice is, the more complicated the structure of the nanodevice is, which limits the speed of DNA assembly. Herein, to address this problem, we investigate the performance of DNA Strand Displacement Reaction (DSDR) in a mixture of polar organic solvents and aqueous buffer and demonstrate that the organic polar solvent can speed up DNA self-assembly efficiently. Taking DSDR in 20% ethanol as an example, first we have demonstrated that the DSDR is highly accelerated in the beginning of the reaction and it can complete 60% of replacement reactions (160% enhancement compared with aqueous buffer) in the first 300 seconds. Secondly, we calculated that the ΔΔG of the DSDR in 20% ethanol (-18.2 kcal mol(-1)) is lower than that in pure aqueous buffer (-32.6 kcal mol(-1)), while the activation energy is lowered by introducing ethanol. Finally, we proved that the DSDR on the electrode surface can also be accelerated using this simple strategy. More importantly, to test the efficacy of this approach in nanodevices with a complicated and slow DNA self-assembly process, we apply this strategy in the hybridization chain reaction (HCR) and prove the acceleration is fairly obvious in 20% ethanol, which demonstrates the feasibility of the proposed strategy in DNA nanotechnology and DNA-based biosensors.

Inspired by biological cell membranes, various "smart" and efficient gating nanoporous devices have been proposed to imitate and to understand life processes. Nanodevices under development with enhanced gating efficiency could play pivotal roles in biosensing and drug delivery. In this Perspective, we highlight an important development by Willner and colleagues that is detailed in this issue of ACS Nano. They designed a new "smart" nanodevice with both "sense" and "release" functionalities for drug delivery based on a nanoporous material, mesoporous silica nanoparticles. We outline recent progress in designing intelligently gated nanoporous devices in material science and nanotechnology. We also summarize new strategies designed for drug delivery based on mesoporous materials. With continuing efforts, we expect more powerful nanodevices to be developed and used in clinical and other real-word applications.

The sensitivity of detection based on biofunctionalized nanopores is limited since the target-to-signal ratio is 1 : 1. Isothermal amplification is a promising amplification strategy at constant temperature due to its easy operation, quick results, PCR-like sensitivity, low cost and energy efficiency. In the present work, the isothermally amplified detection of Zn(2+) is achieved by using a DNA supersandwich structure and Zn(2+)-requiring DNAzymes. The DNA supersandwich structures, due to the multiple amplification of nucleic acids, heavily plug the nanopore. Simultaneously, the DNA supersandwich structures bond with the sessile probe (SP) of the substrate in the nanopore which partially hybridizes with DNAzymes. In the presence of Zn(2+), the Zn(2+)-requiring DNAzyme cleaves the SP into two fragments, while the DNA supersandwich structures are peeled off and the ionic pathway is unimpeded. A steep drop and a sequential complete recovery of the current occur in the I-V plot when the DNA supersandwich structures are decorated and peeled off. In the present system, the reliable detection limit of Zn(2+) is as low as 1 nM. Discrimination between different types of ions (Cu(2+), Hg(2+), Pb(2+)) is achieved.

Potassium is especially crucial in modulating the activity of muscles and nerves whose cells have specialized ion channels for transporting potassium. Normal body function extremely depends on the regulation of potassium concentrations inside the ion channels within a certain range. For life science, undoubtedly, it is significant and challenging to study and imitate these processes happening in living organisms with a convenient artificial system. Here we report a novel biomimetic nanochannel system which has an ion concentration effect that provides a nonlinear response to potassium ion at the concentration ranging from 0 to 1500 microM. This new phenomenon is caused by the G-quadruplex DNA conformational change with a positive correlation with ion concentration. In this work, G-quadruplex DNA was immobilized onto a synthetic nanopore, which undergoes a potassium-responsive conformational change and then induces the change in the effective pore size. The responsive ability of this system can be regulated by the stability of G-quadruplex structure through adjusting potassium concentration. The situation of the grafting G-quadruplex DNA on a single nanopore can closely imitate the in vivo condition because the G-rich telomere overhang is attached to the chromosome. Therefore, this artificial system could promote a potential to conveniently study biomolecule conformational change in confined space by the current measurement, which is significantly different from the nanopore sequencing. Moreover, such a system may also potentially spark further experimental and theoretical efforts to simulate the process of ion transport in living organisms and can be further generalized to other more complicated functional molecules for the exploitation of novel bioinspired intelligent nanopore machines.

Switchable ion channels that are made of membrane proteins play different roles in cellular circuits. Since gating nanopore channels made of proteins can only work in the environment of lipid membrane, they are not fully compatible to the application requirement as a component of those nanodevice systems in which lipid membranes are hard to establish. Here we report a synthetic nanopore-DNA system where single solid-state conical nanopores can be reversibly gated by switching DNA motors immobilized inside the nanopores. High- (on-state) and low- (off-state) conductance states were found within this nanopore-DNA system corresponding to the single-stranded and i-motif structures of the attached DNA motors. The highest gating efficiency indicated as current ratio of on-state versus off-state was found when the length of the attached DNA molecule matched the tip diameter of the nanopore well. This novel nanopore-DNA system, which was gated by collective folding of structured DNA molecules responding to the external stimulus, provided an artificial counterpart of switchable protein-made nanopore channels. The concept of this DNA motor-driven nanopore switch can be used to build novel, biologically inspired nanopore machines with more precisely controlled functions in the near future by replacing the DNA molecules with other functional biomolecules, such as polypeptides or protein enzymes.

Molecular beacon (MB)-based sensing platforms that consist of a fluorogen-quencher pair play an important role in medical and biological researches. However, the synthesis of both fluorogen and quencher in the nucleic acid probes will increase the burden of organic synthesis works and induce the difficulties for precisely controlling the relative distance between fluorogen and quencher, which may lead to false-positive and false-negative results. In this work, initially we report a single labeled MB (FAM-MB, with carboxyfluorescein as fluorogen and without quencher) thus simplifies MBs with the aid of graphene oxide (GO) to detect telomerase activity. To further simplify this structure, namely label-free strategy, we design a facile, sensitive and selective platform using a label-free beacon (AIE-MB, without fluorogen and quencher), based on aggregation-induced emission fluorogen (silole-R). Upon the addition of telomerase, AIE-MB induced comb-like DNA structure leads to high aggregation of silole-R and thus exhibits strong fluorescence emission. By exploitation of this, we can detect telomerase with superior sensitivity and demonstrate their applications in bladder cancer diagnosis. Compared to single-labeled FAM-MB based telomerase activity assay, the label-free AIE-MB induced method could perform the sensitive detection with high signal-to-background ratio.

Almost all of the important functions of DNA are realized by proteins which interact with specific DNA, which actually happens in a limited space. However, most of the studies about the protein–DNA binding are in an unconfined space. Here, we propose a new method, nanopore-based DNA-probe sequence-evolution (NDPSE), which includes up to 6 different DNA-probe systems successively designed in a nanoscale confined space which unveil the more realistic characteristics of protein–DNA binding phenomena. There are several features; for example, first, the edge-hindrance and core-hindrance contribute differently for the binding events, and second, there is an equilibrium between protein–DNA binding and DNA–DNA hybridization.

Herein, we report a novel strategy to accelerate the rate of DNA strand replacement reaction (DSRR) by polar organic solvents. DSRR plays a vital role in DNA nanotechnology but prolonged reaction time limits its further advancement. That is why it is extremely important to speed up the rate of DSRR. In this work, we introduce different polar organic solvents in both simple and complicated DSRR systems and observe that the rate constant is much more than in aqueous buffer. The rate acceleration of DSRR by polar organic solvents is very obvious and we believe that this strategy will extend the application of DNA nanotechnology in future.

Chirality is one of the fundamental biochemical properties in a living system, and a lot of biological and physiological processes are greatly influenced by the chirality of molecules. Inspired by this phenomenon, we study the covalent assembly of DNA on chiral molecule modified surfaces and further discuss the hybridization of DNA on chiral surfaces with nucleic acids. Take methylene blue (MB) modified DNA as a model molecule, we show that the peak current of the L-NIBC (NIBC, N-isobutyryl-L(D)-cysteine) modified gold surface (L-surface) is larger than the D-surface because of a stronger interaction between short-chain DNA and the L-surface; however, the D-surface has a higher hybridization efficiency than the L-surface. Moreover, we apply this result to actual application by choosing an electrochemical DNA (E-DNA) sensor as a potential platform. Furthermore, we further amplify the difference of hybridization efficiency using the supersandwich assay. More importantly, our findings are successfully employed to program the sensitivity and limit of detection.

Recently, the incorporation of biomolecules in Metal-organic frameworks (MOFs) attracts many attentions because of controlling the functions, properties and stability of trapped molecules. Although there are few reports on protein/MOFs composites and their applications, none of DNA/MOFs composite is reported, as far as we know. Here, we report a new composite material which is self-assembled from 3D DNA (guest) and pre-synthesized MOFs (host) by electrostatic interactions and hydrophilic interactions in a well-dispersed fashion. Its biophysical characterization is well analyzed by fluorescence spectroscopy, quartz crystal microbalance (QCM) and transmission electron microscopy (TEM). This new composite material keeps 3D DNA nanostructure more stable than only 3D DNA nanostructure in DI water at room temperature, and stores amounts of genetic information. It will make DNA as a guest for MOFs and MOFs become a new platform for the development of DNA nanotechnology.

Integrating biological components into artificial devices establishes an interface to understand and imitate the superior functionalities of the living systems. One challenge in developing biohybrid nanosystems mimicking the gating function of the biological ion channels is to enhance the gating efficiency of the man-made systems. Herein, we demonstrate a DNA supersandwich and ATP gated nanofluidic device that exhibits high ON-OFF ratios (up to 10(6)) and a perfect electric seal at its closed state (~GΩ). The ON-OFF ratio is distinctly higher than existing chemically modified nanofluidic gating systems. The gigaohm seal is comparable with that required in ion channel electrophysiological recording and some lipid bilayer-coated nanopore sensors. The gating function is implemented by self-assembling DNA supersandwich structures into solid-state nanochannels (open-to-closed) and their disassembly through ATP-DNA binding interactions (closed-to-open). On the basis of the reversible and all-or-none electrochemical switching properties, we further achieve the IMPLICATION logic operations within the nanofluidic structures. The present biohybrid nanofluidic device translates molecular events into electrical signals and indicates a built-in signal amplification mechanism for future nanofluidic biosensing and modular DNA computing on solid-state substrates.

In a traditional sandwich assay, a DNA target hybridizes to a single copy of the signal probe. Here we employ a modified signal probe containing a methylene blue (a redox moiety) label and a "sticky end." When a DNA target hybridizes this signal probe, the sticky end remains free to hybridize another target leading to the creation of a supersandwich structure containing multiple labels. This leads to large signal amplification upon monitoring by voltammetry.

We have demonstrated a novel sensing strategy employing single-stranded probe DNA, unmodified gold nanoparticles, and a positively charged, water-soluble conjugated polyelectrolyte to detect a broad range of targets including nucleic acid (DNA) sequences, proteins, small molecules, and inorganic ions. This nearly “universal” biosensor approach is based on the observation that, while the conjugated polyelectrolyte specifically inhibits the ability of single-stranded DNA to prevent the aggregation of gold-nanoparticles, no such inhibition is observed with double-stranded or otherwise “folded” DNA structures. Colorimetric assays employing this mechanism for the detection of hybridization are sensitive and convenient—picomolar concentrations of target DNA are readily detected with the naked eye, and the sensor works even when challenged with complex sample matrices such as blood serum. Likewise, by employing the binding-induced folding or association of aptamers we have generalized the approach to the specific and convenient detection of proteins, small molecules, and inorganic ions. Finally, this new biosensor approach is quite straightforward and can be completed in minutes without significant equipment or training overhead.

Water-soluble, cationic conjugated polymer binds single-stranded DNA with higher affinity than it binds double-stranded or otherwise "folded" DNA. This stronger binding results from the greater hydrophobicity of single-stranded DNA. Upon reducing the strength of the hydrophobic interactions, the electrostatic attraction becomes the important interaction that regulates the binding between the water-soluble conjugated polymer and DNA. The different affinities between the cationic conjugated polymer and various Forms of DNA (molecular beacons and its open state; single-stranded DNA and double-stranded DNA and single-stranded DNA and complex DNA folds) can be used to design a variety of biosensors.